Fatty acid desaturases of plants have received considerable attention because of their contributions to the physiology and economic value of plants. The activity of fatty acid desaturases, for example, may be a component of the ability of certain species to adjust levels of membrane unsaturation in response to stresses such as chilling (1-3). In addition, the degree of fatty acid unsaturation resulting from desaturase activity is often a major determinant of the nutritional and industrial quality of plant seed oils (4).
Plants typically contain a variety of fatty acid desaturases. The most numerous of these are membrane-associated desaturases that use fatty acids bound to glycerolipids as substrates (5). In addition, the synthesis of oleic acid (18:1.DELTA..sup.9) in plants and certain other organisms such as Euglena (photoauxotrophic) is catalyzed by a desaturase that functions on fatty acids esterified to acyl carrier protein (ACP) (6-8). This enzyme, the .DELTA..sup.9 stearoyl (18:0)-ACP desaturase (EC 1.14.99.6), displays soluble activity in contrast to all previously characterized desaturases (6). In the presence of radiolabeled 18:0-ACP and cofactors including NADPH, ferredoxin (Fd), and ferredoxin-NADPH reductase, the activity of the .DELTA..sup.9 18:0-ACP desaturase is readily detectable in extracts of most plant tissues (9). Due in part to its soluble nature and relative ease of assay, the .DELTA..sup.9 18:0-ACP desaturase has been purified from several plant sources (10-14), and a number of cDNAs encoding this enzyme have been isolated (12-20). In addition to the .DELTA..sup.9 18:0-ACP desaturase, a .DELTA..sup.4 16:0-ACP desaturase has recently been identified in plants (21, 22). This enzyme is a component of the petroselinic acid (18:1.DELTA..sup.6) biosynthetic pathway in endosperm of coriander (Coriandrum sativum L.) and other Umbelliferae species. Translation of a cDNA for the .DELTA..sup.4 16:0-ACP desaturase has revealed that this enzyme shares extensive amino acid identity with the .DELTA..sup.9 18:0-ACP desaturase (21).
The existence of structurally related acyl-ACP desaturases with different substrate recognition and double bond-positioning properties offers the opportunity to compare the active site structures of members of this family of enzymes using techniques such as site-directed mutagenesis and x-ray crystallography. Information gained from this research could potentially lead to the design of desaturases capable of producing new industrially useful isomers of monounsaturated fatty acids. These studies would be aided by the isolation of cDNAs for other variant acyl-ACP desaturases in addition to those for the .DELTA..sup.9 18:0- and .DELTA..sup.4 16:0-ACP desaturases. A potential source of such a desaturase is seed of Thunbergia alata (Acanthaceae family). The oil of this tissue consists of more than 80% weight of the unusual fatty acid .DELTA..sup.6 hexadecenoic acid (16:1.DELTA..sup.6) (23). We have used biochemical and molecular biological approaches to examine whether 16:1.DELTA..sup.6 is synthesized by the activity of a unique acyl-ACP desaturase that is related to the .DELTA..sup.9 18:0- and .DELTA..sup.4 16:0-ACP desaturases.